[Preparation of polyclonal antibody of recombinant human thioredoxin-1 and its protective effects on neonatal rats with endotoxemia].

ABSTRACT

OBJECTIVE: To clone the gene human thioredoxin 1 (hTrx-1) expressing its protein in the E.coli expression system and to obtain its polyclonal antibody, and to study the protective effects of hTrx-1 on neonatal rats with endotoxemia. METHODS: DNA encoding hTrx-1 from fetal liver cells was isolated by RT-PCR. The hTrx-1 was cloned to the prokaryotic expression plasmid PET-28a to induce its protein expression in the E.coli expression system. The purified hTrx-1 was injected into rats to prepare polyclonal antibody. Newborn Sprague-Dawley rats were randomly assigned to three groups control, lipopolysaccharide (LPS) and hTrx-1 (n=12 each). The control and the LPS groups were intraperitoneally injected with normal saline and LPS (5 mg/kg), respectively. The hTrx-1 group received an intraperitoneal injection of hTrx-1 (10 mg/kg) 30 minutes before LPS injection. The mortality rate 24 hrs after injection was compared between the three groups. RESULTS: The prokaryotic expression plasmid PET-28a-hTrx-1 was constructed. The hTrx-1 protein was expressed and purified. The polyclonal antibody of hTrx-1 with the titer of 1∶51200 was prepared. The mortality rate of the control, LPS and hTrx-1 groups was 0, 67% and 17%, respectively (χ2=14.400, P<0.01). CONCLUSIONS: The polyclonal antibody of hTrx-1 is prepared successfully. The hTrx-1 protein has protective effects on neonatal rats with endotoxiamia.